畜牧兽医学报 ›› 2008, Vol. 39 ›› Issue (12): 1748-1752.doi:

• 预防兽医 • 上一篇    下一篇

鸡胚组织膜蛋白与鸡毒霉形体黏附蛋白的结合分析

彭秀丽1,2,高俊伟1,胡思顺1,胡福利1,毕丁仁1*   

  1. 1.华中农业大学动物医学院,武汉 430070;2.华中农业大学动物遗传育种与繁殖教育部重点实验室,武汉 430070
  • 收稿日期:1900-01-01 修回日期:1900-01-01 出版日期:2008-12-24 发布日期:2008-12-24
  • 通讯作者: 毕丁仁

Binding Research Between the Tissues Membrane of Chick Embryo and pMGA

PENG Xiu-li1,2,GAO Jun-wei1,HU Si-shun1,HU Fu-li1,BI Ding-ren1*   

  1. 1.College of Veterinary Medicine,Huazhong Agricultural University,Wuhan 430070,China 2.Key Laboratory of Agricultural Animal Genetics,Breeding and Reproduction of Ministry of Education, Huazhong Agricultural University, Wuhan 430070,China
  • Received:1900-01-01 Revised:1900-01-01 Online:2008-12-24 Published:2008-12-24

摘要: 利用pGEX融合蛋白表达系统,将鸡毒霉形体黏附蛋白(pMGA)与GST的编码序列在大肠杆菌BL21中进行融合表达,经 GST·Bind Resin 纯化,GST-pMGA的纯度达96%。用蛋白酶Thrombin切掉GST标签,获得纯度为96%的pMGA,经Western-blot鉴定具有良好的免疫学活性。通过差速离心法提取SPF鸡胚的气管、心脏、肝脏、肺脏、肾脏、腺胃、十二指肠、法氏囊、胸腺和脾脏等组织膜蛋白,采用斑点酶联免疫吸附试验(Dot-ELISA)检测各组织膜蛋白与pMGA的结合。结果表明:除脾脏外,其余组织的膜蛋白与pMGA之间存在特异性结合,说明这些组织膜蛋白中存在pMGA的受体蛋白。经SDS-PAGE分析发现,除脾脏外,其余9种组织的膜蛋白中均含有一条相对分子质量约为30 ku的主带,受体的相对分子质量可能为30 ku,可为深入研究pMGA的受体蛋白提供资料。

Abstract: The GST-pMGA was expressed in E. coli BL21 with the pGEX fusion expression system. GST-pMGA expressed in supernatant was purified using GST Bind Resin and a plenty of soluble protein was obtained, the final preparation was 96% after purification by SDS-PAGE analysis. GST-pMGA was incubated with thrombin protease overnight, the pMGA was obtained and showed well immunological activity by Western-blot analysis. The tissues membrane protein from trachea, heart, lung, liver, kidney, thymus, bursa of Fabricius, spleen, proventriculus and the duodenum of SPF chick embryo were obtained through centrifugation, Dot-ELISA was applied to detect the specific binding relation between the membrane protein and pMGA. The results showed that pMGA and the tissues membrane protein of chick embryo had a specific binding except the spleen, this binding indicated receptor of pMGA on the tissues membrane protein of chick embryo. In membrane protein extracted from these tissues except spleen, there are the same product with Molecular weight approximately 30 kD by SDSPAGE analysis, so it was concluded it’s the possible protein interacting with pMGA. All this makes it possible to do further studies of receptor of pMGA.